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251 Furthermore, both obesity and aging drive the expansion of double-negative (DN) B cells (CD27IgD), which display a proinflammatory phenotype marked by autoimmune antibody secretion and upregulated expression of activation markers (e.g., CD11c and T-bet)

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For the IHC study, after deparaffinization (60 C) and antigen retrieval (1 mM citrate, pH 6, 98 C), slides were treated with 3% H 2 O 2 (5 min) and incubated with primary antibodies (1550 min): cleaved caspase-3 (1:300 dilution, Cell Signaling, 9664S), -H2AX (1:400 dilution, Abcam ab22551), HO-1 (1:400 dilution, Abcam ab52947) and CDA (1:300 dilution, Invitrogen PA5-95616) on a Leica Bond RXm autostainer